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Application of berberine in medicament for treating hepatitis B virus infection

A technology of hepatitis B virus and berberine, applied in the field of medicine, can solve problems such as the severe situation of hepatitis B prevention and treatment, and achieve the effects of improving treatment quality and medication compliance, abundant drug resources, and convenient use.

Inactive Publication Date: 2010-02-10
WUHAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although my country has implemented hepatitis B preventive vaccination, the situation of hepatitis B prevention and control is severe due to the large number of virus carriers, immune tolerance, drug resistance, the infected person is the source of life-long virus infection, and the virus is transmitted from mother to child.

Method used

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  • Application of berberine in medicament for treating hepatitis B virus infection
  • Application of berberine in medicament for treating hepatitis B virus infection
  • Application of berberine in medicament for treating hepatitis B virus infection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1: Cytotoxicity of berberine to HepG2 cells and HepG2 2.15 cells

[0037] Disperse well-grown HepG2 cells and HepG2 2.15 cells into a single cell suspension with trypsin, press 1×10 5 / ml concentration in 96-well plate, 0.1ml per well, set at 37°C, 5% CO 2 Cultivate in the incubator for 24h. After the cells grow into a single layer, the supernatant of the culture medium is discarded and replaced with a drug-containing maintenance solution of different concentrations. Three replicate wells are set for each concentration, and a normal cell control is set. Continue culturing for 48 h, discard the culture supernatant, add 100 μl of serum-free DMEM medium containing 100 ug / ml of MTT (tetramethylazozolium salt) to each well, and place in CO 2 After continuing to culture in the incubator for 2-3 hours, discard the MTT supernatant, wash 3 times with PBS, add 100 μl of dissolving solution (DMSO:ethanol volume ratio: 1:1) to each well, shake for 5-10 minutes, and set th...

Embodiment 2

[0039] The results showed that the drug had no obvious cytotoxicity in the concentration range of 0.32ug / ml-1000ug / ml. Example 2: Effect of berberine on HBsAg and HBeAg in HepG2 2.15 cells in vitro

[0040] 1) Take the coated microwell reaction strip, fix it on the plastic frame, add 50 μl / well of the sample, 50 μl (or 1 drop) / well of the negative control and 50 μl (or 1 drop) of the sex control in each two wells, and leave one well without adding it as a blank control, and then add Measure HBsAg or HBeAg enzyme conjugate 50μl (or one drop) / well, blank control is not added.

[0041] 2) Place a constant temperature water bath at 37°C for 30 minutes.

[0042] 3) Carefully remove the liquid in the wells and pat dry; dilute the washing solution 1:20 times with distilled water, fill each well, and discard after staying for 30 seconds. Repeat this way for 6 times and pat dry each time. Avoid cross-contamination between wells as much as possible.

[0043] 4) Add 50 μl (or one drop...

Embodiment 3

[0061] Example 3: Effects of Drugs on Some Classical Signaling Pathways

[0062] 1. Lipofectamine transfection method:

[0063] The negatively charged DNA binds to the positively charged end of the cationic compound, and then the DNA-binding lipid binds to the cell membrane, allowing the DNA to enter the cell. The liposome transfection method can produce extremely high transfection efficiency for some cells. The liver cells HepG2 used in this experiment (purchased from the China Typical Microorganism Collection Center) have been confirmed by long-term transfection experiments in the laboratory. Dyeing is better. The specific method takes the transfection operation as an example with a 24-well plate. The whole process of transfection must be strictly aseptic.

[0064] 1) Seed the cells to be transfected in a 24-well plate one day before transfection and culture at 37°C. The cell density before transfection was 40-60%.

[0065] 2) Prepare the following solution

[0066] a)...

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Abstract

The invention discloses the application of berberine in medicament for treating hepatitis B virus infection. In the invention, adopting an HepG2.2.15 cell model for the in vitro selection of anti-hepatitis B virus medicine, preliminarily selecting the effect of the medicament by measuring the difference of HBeAg and HBsAg expression volumn in supernatant fluid of the cell after administration, andfurther determining the effectiveness of the medicine by studying the inhibitory effect of the HBV DNA reproduction level (ccc DNA) in the HepG2.2.15 cell by the medicament. Meanwhile, the influenceon the reproduction of the hepatitis B virus by the transcriptional level of the medicine is studied according to the activity inhibitory effect to HBV viral promoters by the medicine, and key HBV cis-regulatory elements sensitive to the antivirus medicine are found. A pivotal cellular signal transduction pathway possibly participating the transcription reproduction for inhibiting viruses by the medicine is analyzed and verified on the basis, the antivirus molecular mechanism of the medicament is further studied, and the medicament is indicated to be developed and applied to clinical treatmentas an anti-hepatitis B effective medicament.

Description

technical field [0001] The invention relates to the technical field of medicine, in particular to the application of berberine in the preparation of medicines for treating or preventing hepatitis B virus infection. Background technique [0002] HBV is the prototype virus of the Hepadanaviridae family. The HBV genome is an incomplete double-stranded circular DNA with a length of about 3200 base pairs (kb). It has the following characteristics: ① Each ORF region in the negative strand of HBV DNA is a viral antigen protein coding sequence, and the cis-regulatory elements in the genome are all located in each coding region, including 4 promoters and 2 enhancers. The three promoters regulate the mRNAs of genome C and PreC, L protein, M protein, S protein, and X protein respectively. Envelope protein S is a protein encoded by the S gene (hepatitis B surface antigen, HBsAg). HBsAg is associated with viral adsorption to receptors on the surface of liver cells, which can alter the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/4375A61P31/20
Inventor 吴建国陶君彦朱应庞然袁婷婷
Owner WUHAN UNIV
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